Lymphocyte Activation Gene-3, or LAG-3 (also known as Cluster of Differentiation 223 or CD223) is a membrane protein of the immunoglobulin supergene family. LAG-3 is structurally and genetically related to CD4, with its encoding gene located on the distal part of the short arm of chromosome 12, near the CD4 gene, suggesting that the LAG-3 gene may have evolved through gene duplication (Triebel et al., J Exp Med, 1990). LAG-3 is not expressed on resting peripheral blood lymphocytes but is expressed on activated T cells and natural killer (NK) cells (Triebel et al., J Exp Med, 1990), and has been reported to also be expressed on activated B cells (Kisielow et al., Eur J Immunol, 2005) and plasmacytoid dendritic cells (Workman et al., J Immunol, 2009).
Like CD4, LAG-3 binds to major histocompatibility complex (MHC) class II molecules, but with a two-fold higher affinity and at a different binding site (Huard et al., Proc Natl Acad Sci USA, 1997). MHC class II engagement on dendritic cells by LAG-3 leads to changes in the cytokine and chemokine profiles of dendritic cells (Buisson and Triebel, Vaccine, 2003), Further, LAG-3 has been reported to cause maturation of dendritic cells, as demonstrated by the production of IL-12 and TNF-alpha by these cells and increases in the capacity of dendritic cells to stimulate the proliferation and IFN-gamma response by allogeneic T-cells (Andreae et al., J Immunol, 2002). LAG-3 signaling and MHC class II cross-linking has been reported to inhibit early events in primary activation of human CD4+ and CD8+ T-cells (Macon-Lemaitre and Triebel, Immunology, 2005). It negatively regulates the cellular proliferation, activation and homeostasis of T cells.
Like CTLA-4 and PD-1, LAG-3 is an inhibitory immune receptor. LAG-3's prominent role as a negative regulator of T cell responses has been impressively demonstrated in particular in conjunction with PD-1 in a study based on both knockout mice and target-specific antibodies (Woo et al., Cancer Res, 2012). In this study, dual anti-LAG-3/anti-PD-1 antibody treatment cured most mice of established tumors that were largely resistant to single antibody treatment. Further, LAG-3/PD-1 double knock-out mice showed markedly increased survival from and clearance of multiple transplantable tumors. Further strong experimental support for the powerful combined role of PD-1 and LAG-3 as immune checkpoints was provided by the fact that the double knock-out mice were highly prone to lethal autoinflammation.
Consequently, there exists an unmet need in the art for compounds that modulate responses of LAG-3+ lymphocytes, such as T-cells, NK cells, B cells, and plasmacytoid dendritic cells, which may have important uses in the treatment or prevention of cancer, organ transplant rejection, or treatment of autoimmune or autoinflammatory diseases. In this regard, the present disclosure provides a group of novel compounds specifically binding to LAG-3, thereby, modulating the immune response. Such compounds are muteins derived from lipocalins. Muteins of lipocalins are a rapidly expanding class of therapeutics and can be constructed to exhibit high affinity and specificity against desired targets (see e.g. International Patent Publication Nos, WO 99/16873, WO 00/75398, WO 03/029471, WO 05/19256).